This proposal explores aspects of the molecular mechanisms regulating subcellular distribution of the thyroid hormone receptor (TR), a transcription factor which either activates or represses the expression of its target genes in response to thyroid hormone (T3). TR is essential for normal development, growth, and homeostasis. Rapid shuttling of TR between the nuclear and cytoplasmic compartments may provide an additional checkpoint in the control of T3-responsive gene expression. Whether nuclear import/retention signals are dominant to nuclear export signals in TR, and whether the rapidity and extent of TR shuttling is ligand-dependent will be tested, by analyzing the subcellular trafficking of green fluorescent protein-tagged TR in mammalian cell heterokaryons. Mutant receptors are associated with a number of endocrine and neoplastic diseases. Several of these dominant negative variants of TR, including the oncoprotein v-ErbA, have distinct subcellular distribution patterns. Surprisingly, although TR nuclear export does not follow a pathway mediated by the export receptor CRM1, v-ErbA export is CRM1-dependent. Amino acid sequences that differ between v-ErbA and TR will be examined to determine which mutations in v-ErbA confer CRM1-dependence on the oncoprotein's nuclear export pathway. Whether the availability of intranuclear binding sites is a key determinant for nuclear retention of TR will be tested, by investigating the role of ligand binding and heterodimer formation in partitioning of TR to the nuclear matrix. The functional significance of matrix association for transcriptional activation and silencing will be assessed, by determining whether dominant negative variants of TR associate with the nuclear matrix. These studies will not only increase understanding of the normal cellular response to T3, but should also provide important insight into the ontogeny of an oncogene and modulation of gene expression through both compartmentalization and dominant negative transcription factors.